Immunological characterization of various Pseudomonas pseudomallei preparations was carried out by SDS-PAGE and Western blot using sera from infected humans and from patients with other bacterial infections. Somatic (SOM) and partially purified cell extracts (PCE) gave more complex SDS-PAGE patterns: M(r) ranged from 86 to 12.7 and 48 to 10 kDa, respectively. The culture-filtrated antigens (CF) from 3 different kinds of synthetic media consisted of fairly simple profiles with common bands M(r) of 40, 26 and 16 kDa. PCE and CF reacted specifically with infected human sera; SOM did not. The components with M(r) of 40 kDa in CF reacted consistently with all infected sera but failed to react with sera infected with Escherichia coli, Enterobacter spp., Klebsiella pneumoniae, Proteus mirabilis, Salmonella spp., Staphylococcus aureus, Streptococcus spp., Pseudomonas aeruginosa and P. stutzeri. This peptide was demonstrated to be a major component in CF thus suggesting its potential for development of immunodiagnostic methods for melioidosis.